This study will investigate a novel gene delivery system that is based on the formation of sulfhydryl crosslinks within peptide DNA condensates. The gene delivery system under investigation is a multi-component peptide condensed DNA formulation. The key components consist of plasmid DNA condensed by an addmixture of peptide conjugates each containing multiple sulfhydryl groups designed to spontaneously polymerize and cross-link when bound to DNA. Following i.v. dosing, cross-linked DNA condensates are proposed to transiently stabilize plasmid DNA during circulation and inside the target cells. Cross-linking peptides will be covalently derivatized with a single polyethylene glycol (PEG) chain to form a steric layer on the surface of DNA condensates that blocks protein binding and masks DNA condensate recognition by the reticuloendothelial system. Targeting specificity will be achieved by derivatizing a cross-linking peptide with a single N-glycan resulting in glycopeptides that direct targeting to either the asialoglycoprotein receptor on hepatocytes or the mannose receptor on Kupffer cells. DNA condensates will be prepared using binary addmixtures of cross-linking glycopeptide and PEG-peptide. Cross-linking peptides will be modified to buffer endosomes and allow DNA condensates to release into the cytosol of target cells. Once in the cytosol, crosslinked DNA condensates are proposed to slowly release plasmid DNA following disulfide bond reduction. The central hypothesis to be tested is that increasing DNA condensate stability and controlling the rate of DNA release into the cytosol will prolong the half-life of DNA and produce higher levels of gene expression in vivo. These studies aim to systematically optimize the level of transient gene expression in vivo using a novel chemical mechanism to overcome barriers that currently limit the efficiency of nonviral gene delivery systems.